The biogenic synthesis of silver nanoparticles (AgNPs) has attracted considerable attention for biomedical applications due to their eco-friendly nature and the use of readily available plant extracts. In this study, green-synthesised AgNPs from Senna didymobotrya leaves aqueous extract were evaluated for their physicochemical characteristics, stability, and in vitro cytotoxic and phototoxic activity in comparison with chemically synthesised AgNPs. Nanoparticle formation was confirmed by distinct colour changes, with surface plasmon resonance peaks at 401 nm and 466 nm for chemically and green-synthesised AgNPs, respectively. FTIR analysis indicated bioactive compounds capping on green AgNPs, whereas chemical AgNPs showed citrate-associated signals. DLS and zeta potential measurements demonstrated stable dispersions with sizes of 96.9 nm (–26.3 mV) for green AgNPs and 80.3 nm (–30.8 mV) for chemical AgNPs. TEM and SEM confirmed spherical morphology, with smaller, aggregated chemical AgNPs (24 ± 8.1 nm) and larger, well-capped green AgNPs (39 ± 8.1 nm). Under laser irradiation of chemical AgNPs (405 nm) and green AgNPs (470 nm), chemical AgNPs showed moderate photothermal-induced temperature increases (2.8 °C), compared to negligible temperature changes (0.8 °C), like the water control (0.9 °C), for green AgNPs. However, the photochemical generation of reactive oxygen species was 7-fold higher for green AgNPs. The MTT assay results revealed comparatively enhanced chemo and phototoxic activity of green AgNPs against MCF-7 and MDA-MB-231, mediated predominantly by apoptosis, as supported by Annexin V/PI analysis distinguishing early and late apoptotic populations. Overall, findings suggest biosynthesised AgNPs are promising anticancer agents due to synergistic chemo-phototoxic efficacy.